PROJECT SUMMARY To efficiently multiply in their hosts, viruses must hijack the host protein synthesis machinery and overcome potent cell-intrinsic, anti-viral immune responses. A powerful common strategy utilized by many different patho- genic human viruses to achieve these ends involves accelerating global mRNA decay. Until recently, this has been regarded primarily as a means to restrict production of host anti-viral proteins, promote translation of abundant viral mRNAs, and facilitate transitions between different temporal classes of viral mRNAs. In cells infected with the prototypical poxvirus Vaccinia, global mRNA decay is stimulated by viral decapping enzymes that presumably generate substrates for degradation by the cellular 5'-3' mRNA exonuclease Xrn1. Unexpect- edly, our preliminary results demonstrated that the host Xrn1 is required to limit accumulation of double- stranded RNA (dsRNA), a pathogen-associated molecular pattern (PAMP) that activates host anti-viral defen- ses and results in inhibition of protein synthesis and viral replication. Moreover, Xrn1-depletion sensitized uninfected cells to exogenous dsRNA, excluding the possibility that the response is specific to Vaccinia- encoded dsRNA. This raises the exciting possibility that other viruses liable to generate dsRNA may rely on Xrn1 function to avoid immune detection. Our overall objective is to understand how cell intrinsic antiviral immunity is controlled by host RNA decay enzymes. Unlike Vaccinia virus, herpes simplex virus-1 (HSV1) replicates in the nucleus and does not encode decapping enzymes. Importantly, cytoplasmic dsRNA reportedly accumulates in HSV1-infected cells and global mRNA decay is accelerated via the virus-encoded vhs mRNA endonuclease. Endonucleolytic mRNA cleavage by vhs produces substrates with 5'-monophosphate termini that are substrates for Xrn1, although the role of Xrn1 in HSV1 infection biology remains largely unknown. Based on our preliminary data, we hypothesize that the host mRNA decay enzyme Xrn1 regulates cytoplasmic dsRNA accumulation in HSV1-infected cells. Here, this hypothesis is tested in two specific aims that i) deter- mine the role of the 5'-3' mRNA exonuclease Xrn1 in HSV-1 infection biology; and ii) define how the HSV1 mRNA endonuclease vhs collaborates with Xrn1 to regulate dsRNA abundance and anti-viral immune respons- es. Results from these experiments will impact our understanding of how host mRNA decay enzymes regulate cell intrinsic anti-viral immunity in cells infected with the human herpesvirus HSV1. Furthermore, investigating how Xrn1 regulates dsRNA accumulation and anti-viral immunity are potentially applicable to a wide variety of pathogenic human viruses and could reveal new strategies for treating virus infections. Finally, this study will provide insight into how cellular mRNA decay enzymes regulate nucleic acid-responsive, cell intrinsic immune pathways and have significant implications for understanding both autoimmune disease and infection biology.